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Regulation of environmental r esponses in the Pacific oyster, Crassostrea gigas

Regulation of environmental r esponses in the Pacific oyster, Crassostrea gigas. Mackenzie Gavery University of Washington School of Aquatic and Fishery Sciences Committee Meeting: April 15 th , 2014. Agenda:. Research update Proposed t imeline. Environmental Response in Oysters.

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Regulation of environmental r esponses in the Pacific oyster, Crassostrea gigas

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  1. Regulation of environmental responses in the Pacific oyster, Crassostreagigas Mackenzie Gavery University of Washington School of Aquatic and Fishery Sciences Committee Meeting: April 15th, 2014

  2. Agenda: • Research update • Proposed timeline

  3. Environmental Response in Oysters

  4. Environmental Response in Oysters Cyclooxygenase characterization

  5. Environmental Response in Oysters Cyclooxygenase characterization Gene expression: disease resistance

  6. Environmental Response in Oysters Chapter I. CBP: Part D Cyclooxygenase characterization Whole transcriptome Gene expression: disease resistance

  7. Chapter I. RNA-Seq LOW IMPACT HIGH IMPACT

  8. Chapter I. RNA-Seq LOW IMPACT HIGH IMPACT 742 novel contigs 427 differentially expressed

  9. Chapter I. RNA-Seq LOW IMPACT HIGH IMPACT 742 novel contigs 427 differentially expressed Ultra-short read technology is a powerful tool for gene discovery and expression analysis in organisms with limited genomic resources

  10. Environmental Response in Oysters Chapter I. CBP: Part D Cyclooxygenase characterization Whole transcriptome Gene expression: disease resistance

  11. Environmental Response in Oysters Chapter I. CBP: Part D Cyclooxygenase characterization Whole transcriptome Gene expression: disease resistance DNA methylation: single gene/in silico Chapter II. BMC Genomics Frontiers

  12. Chapter II. in silicoanalysis ‘inducible’ ‘housekeeping’ Predicted degree of DNA methylation

  13. Chapter II. in silico analysis DNA methylation detected by targeted bisulfite sequencing and methylation specific PCR DNA methylation patterns are different between functionally distinct classes of genes ‘inducible’ ‘housekeeping’ Predicted degree of DNA methylation

  14. Environmental Response in Oysters Chapter I. CBP: Part D Cyclooxygenase characterization Whole transcriptome Gene expression: disease resistance DNA methylation: single gene/in silico Chapter II. BMC Genomics Frontiers

  15. Environmental Response in Oysters Chapter I. CBP: Part D Cyclooxygenase characterization Whole transcriptome Gene expression: disease resistance DNA methylation: single gene/in silico Nanostring Chapter II. BMC Genomics Frontiers

  16. Environmental Response in Oysters Chapter I. CBP: Part D Cyclooxygenase characterization Whole transcriptome transgenerational (5-aza pesticide) EE2 Gene expression: disease resistance DNA methylation: single gene/in silico Nanostring Chapter II. BMC Genomics Frontiers

  17. Environmental Response in Oysters Chapter I. CBP: Part D Cyclooxygenase characterization Whole transcriptome transgenerational (5-aza pesticide) EE2 bringing it all together! meth & expression Gene expression: disease resistance DNA methylation: single gene/in silico Nanostring Chapter II. BMC Genomics Frontiers

  18. Environmental Response in Oysters Chapter I. CBP: Part D Cyclooxygenase characterization Whole transcriptome Ch III. PeerJ transgenerational (5-aza pesticide) EE2 bringing it all together! meth & expression Gene expression: disease resistance DNA methylation: single gene/in silico Nanostring Chapter II. BMC Genomics Frontiers

  19. Environmental Response in Oysters Chapter I. CBP: Part D Chapter IV. Brief Func Gen Cyclooxygenase characterization Whole transcriptome Ch III. PeerJ transgenerational (5-aza pesticide) EE2 bringing it all together! meth & expression Gene expression: disease resistance DNA methylation: single gene/in silico Nanostring Chapter II. BMC Genomics Frontiers

  20. Chapter III: whole methylome • Approach • High-throughput bisulfite sequencing genomic DNA

  21. Chapter III: whole methylome • Approach • High-throughput bisulfite sequencing genomic DNA

  22. Chapter III: whole methylome • Approach • High-throughput bisulfite sequencing • RNA-Seq data genomic DNA

  23. Chapter III: whole methylome • Gill tissue: > 2.5 million CG dinucleotides

  24. Chapter III: whole methylome 0bp 200,000bp CG genes exons ex %methylation 100% 0% scaffold 86 (Galaxy Trackster)

  25. Chapter III: whole methylome 0bp 200,000bp CG genes exons ex %methylation 100% 0% scaffold 86 (Galaxy Trackster)

  26. Chapter III: whole methylome 0bp 200,000bp CG genes exons ex %methylation 100% 0% scaffold 86 (Galaxy Trackster)

  27. Chapter III: whole methylome 0bp 200,000bp CG genes exons ex %methylation 100% 0% scaffold 86 (Galaxy Trackster)

  28. Chapter III: whole methylome • Distribution in genomic elements

  29. Chapter III: whole methylome • Distribution in genomic elements

  30. Chapter III: whole methylome • Relationship with expression

  31. Chapter III: whole methylome • Relationship with expression DNA methylation/gene Gene expression (Deciles) RNA-Seq data (Zhang et al., 2012)

  32. Chapter III: whole methylome • Relationships with other gene specific attributes?

  33. Chapter III: whole methylome • Relationships with other gene specific attributes? • Spatial • e.g. number of exons, length of mRNA • Gene expression (Zhang et al 2012): • Mean expression level across 9 tissues • Variation in expression between tissues (%CV)

  34. Chapter III: whole methylome PC2 (26.1%) PC1 (50.2%)

  35. Chapter III: whole methylome PC2 (26.1%) PC1 (50.2%)

  36. Chapter III: summary

  37. Chapter III: summary methylated unmethylated

  38. Chapter III: summary methylated unmethylated Gene function:

  39. Chapter III: summary methylated unmethylated Gene function: inducible housekeeping

  40. Chapter III: summary methylated unmethylated Gene function: inducible housekeeping Expression:

  41. Chapter III: summary methylated unmethylated Gene function: inducible housekeeping low high Expression:

  42. Chapter III: summary methylated unmethylated Gene function: inducible housekeeping low high Expression: Tissue specific expression:

  43. Chapter III: summary methylated unmethylated Gene function: inducible housekeeping low high Expression: Tissue specific expression: variable conserved

  44. Chapter III: summary methylated unmethylated Gene function: inducible housekeeping low high Expression: Tissue specific expression: variable conserved Chapter IV. A context dependent role of DNA methylation in bivalves (Gavery & Roberts 2014)

  45. Summary of Chapters: • Transcriptomic approaches to understanding environmental response • First characterization of DNA methylation in oysters (in silicoanalysis) • DNA methylation & gene expression: whole genome analysis • Perspective/Review: DNA methylation in bivalves

  46. Environmental Response in Oysters Chapter I. CBP: Part D Chapter IV. Brief Func Gen Cyclooxygenase characterization Whole transcriptome Ch III. PeerJ transgenerational (5-aza pesticide) EE2 bringing it all together! meth & expression Gene expression: disease resistance DNA methylation: single gene/in silico Nanostring Chapter II. BMC Genomics Frontiers

  47. Environmental Response in Oysters Chapter I. CBP: Part D Chapter IV. Brief Func Gen Cyclooxygenase characterization Whole transcriptome Ch III. PeerJ transgenerational (5-aza pesticide) EE2 bringing it all together! meth & expression Gene expression: disease resistance DNA methylation: single gene/in silico Nanostring BMC Genomics Frontiers Authorea link

  48. Agenda: • Research update • Proposed timeline

  49. Proposed Timeline: • Draft proposal to reading committee: April 21st

  50. Proposed Timeline: Spring End Mem Day

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